Figure 4. TRPV1-mediated Ca²⁺ influx was required to maintain DPSC function.

a DPSC proliferation was increased after TRPV1 knockdown, as assessed by BrdU proliferation assay. b, c DPSC osteogenic/dentinogenic differentiation was decreased after TRPV1 knockdown, as assessed by Alizarin red staining, with decreased expression of RUNX2, ALP, and DSPP analyzed by qPCR. d Bone/dentin-like hard tissue formation of DPSCs in vivo decreased after TRPV1 knockdown by siRNA. e ALP-positive and DSPP-positive cells detected in tissues formed by DPSC transplantation were decreased after TRPV1 knockdown. f NaHS treatment upregulated p-GSK3β expression, and knockdown of TRPV1 by siRNA partially blocked NaHS-induced GSK3β phosphorylation. g CBS or CSE siRNA treatment downregulated p-GSK3β expression. h H₂S donor NaHS treatment activated β-catenin expression, which was partially blocked after TRPV1 knockdown. i NaHS treatment enhanced TOPflash activity in DPSCs, which was blocked after TRPV1 siRNA treatment, as assessed by luciferase activity. NaHS treatment failed to alter FOPflash activity. HA HA/TCP, CT connect tissue, B/D bone/dentin. *P < 0.05, **P < 0.01; scale bar: 100 μm (b, d), 50 μm (e). All experimental data were verified in at least three independent experiments